Each year about 50,000 new cases of ductal carcinoma in situ (DCIS) are diagnosed in the US, and account for 17-34% of all mammographically detected cases. Disease recurs within 10 years in 20% of women treated for DCIS. Currently, no clinical or histopathological characteristics have been found that are consistently associated with recurrence. Promoter hypermethylation is a powerful epigenetic mechanism for suppression of gene expression and occurs in many types of cancers. Hypermethylated genes provide very sensitive and specific DMA markers for breast cancer. A quantitative method [Quantitative Multiplex- Methylation Specific PCR (QM-MSP)] was developed in the investigator's laboratory, which detects hypermethylation in multiple genes in very small biological samples of cancer cells in the presence of a vast excess of normal cells, with high sensitivity and specificity. This proposal is based upon the hypothesis that the hypermethylation of a specific set of genes in DCIS can be used to recognize those DCIS cases that will recur as invasive cancer. The goal of this proposal is to identify genes that are methylated in DCIS, study their behavior during tumor progression, and develop a recurrence model based on a DCISspecific gene methylation panel to test their utility as predictors of tumor recurrence in patients with long-term follow-up This goal will be achieved in three steps: In Aim 1, perform a genome-wide scan for genes silenced by hypermethylation using a modified array analysis on purified epithelial cells, isolated from 30 primary DCIS and treated with demethylating agents. In Aim 2, determine if the genes identified in Aim 1 support a model of progression from DCIS to invasive breast cancer. The genes, following verification, will be used for QM-MSP analysis of archival normal, benign, DCIS and invasive cancer samples, and in analogous samples obtained from 44 individual mastectomy specimens. In Aim 3, test the ability of the methylated gene marker panel derived in Aim 2 to prognosticate recurrence of DCIS as invasive ductal carcinoma, and thereby identify patients at risk. The most promising gene panel will be tested in training and test sets by QM-MSP in tumor samples from UCSF's DCIS case-control study. In this cohort of 1062 women treated only by lumpectomy, 200 cases recurred within 7 years, and are matched with 400 age-, grade-, size- and marginmatched control cases that did not recur. As a result of this work, we hope to derive an algorithm that will identify DCIS patients at risk for recurrence, and derive a molecular progression paradigm that will permit the development of a therapeutic strategy that will improve tumor-free survival among those patients, while minimizing excessive therapy in those unlikely to develop invasive disease.